I. Field of Invention
This invention relates to a method of using alpha-methylglucoside (AMG) in vivo as an indicator for glucose absorption from the gastrointestinal (GI) system, glucose re-absorption from the kidney tubules, and/or glucose excretion in the urine, after oral administration of AMG.
II. Description of Related Art
Hyperglycemia, that is, elevated plasma glucose, is a hallmark of diabetes. Type I diabetes mellitus, which comprises approximately 10% of diabetes cases, was previously referred to as insulin-dependent diabetes mellitus (“IDDM”) or juvenile-onset diabetes. This disease is characterized by a progressive loss of insulin secretory function by beta cells of the pancreas. This characteristic is also shared by non-idiopathic, or “secondary”, diabetes having its origins in pancreatic disease. Type I diabetes mellitus is associated with the following clinical signs or symptoms: persistently elevated plasma glucose concentration or hyperglycemia; polyuria; polydipsia and/or hyperphagia; chronic microvascular complications such as retinopathy, nephropathy and neuropathy; and macrovascular complications such as hyperlipidemia and hypertension which can lead to blindness, end-stage renal disease, limb amputation and myocardial infarction.
Type II diabetes mellitus (non-insulin-dependent diabetes mellitus or NIDDM) is a metabolic disorder involving the dysregulation of glucose metabolism and impaired insulin sensitivity. Type II diabetes mellitus usually develops in adulthood and is associated with the body's inability to utilize or make sufficient insulin. In addition to the insulin resistance observed in the target tissues, patients suffering from type II diabetes mellitus have a relative insulin deficiency, with lower than predicted insulin levels for a given plasma glucose concentration. Type II diabetes mellitus is characterized by the following clinical signs or symptoms: persistently elevated plasma glucose concentration or hyperglycemia; polyuria; polydipsia and/or hyperphagia; chronic microvascular complications such as retinopathy, nephropathy and neuropathy; and macrovascular complications such as hyperlipidemia and hypertension which can lead to blindness, end-stage renal disease, limb amputation and myocardial infarction.
Syndrome X, also termed Insulin Resistance Syndrome (IRS), Metabolic Syndrome, or Metabolic Syndrome X, is recognized in some 2% of diagnostic coronary catheterizations. Often disabling, it presents symptoms or risk factors for the development of Type II diabetes mellitus and cardiovascular disease, including impaired glucose tolerance (IGT), impaired fasting glucose (IFG), hyperglycemia, hyperinsulinemia, insulin resistance, dyslipidemia (e.g., high triglycerides, low HDL), hypertension and obesity.
Therapy for IDDM patients has consistently focused on administration of exogenous insulin, which may be derived from various sources (e.g., human, bovine, porcine insulin). The use of heterologous species material gives rise to formation of anti-insulin antibodies which have activity-limiting effects and result in progressive requirements for larger doses in order to achieve desired hypoglycemic effects.
Typical treatment of Type II diabetes mellitus focuses on maintaining the blood glucose level as near to normal as possible with lifestyle modification relating to diet and exercise, and when necessary, the treatment with anti-diabetic agents, or insulin, or a combination thereof. First-line therapy for NIDDM that cannot be controlled by dietary management is treatment with oral antidiabetic agents.
First-line therapies for NIDDM typically include metformin and sulfonylureas as well as thiazolidinediones. Metformin monotherapy is a first line choice, particularly for treating type II diabetic patients who are also obese and/or dyslipidemic. Lack of an appropriate response to metformin is often followed by treatment with metformin in combination with sulfonylureas, thiazolidinediones, or insulin. Sulfonylurea monotherapy (including all generations of drugs) is also a common first line treatment option. Another first line therapy choice may be thiazolidinediones. Alpha glucosidase inhibitors are also used as first and second line therapies. Patients who do not respond appropriately to oral anti-diabetic monotherapy are given combinations of the above-mentioned agents. When glycemic control cannot be maintained with oral antidiabetics alone, insulin therapy is used either as a monotherapy or in combination with oral antidiabetic agents.
Although insulin resistance is not always treated in all Syndrome X patients, those who exhibit a prediabetic state (e.g., IGT, IFG), where fasting glucose levels may be higher than normal but not at the diabetes diagnostic criterion, are treated in some countries (e.g., Germany) with metformin in an effort to prevent diabetes. The anti-diabetic agents may also be combined with other pharmacological agents for the treatment of the concomitant co-morbidities (e.g., antihypertensives for hypertension, hypolipidemic agents for lipidemia).
A recent development in treating hyperglycemia is focused on excretion of excessive glucose directly into the urine. Plasma glucose is normally filtered in the kidney in the glomerulus and actively reabsorbed in the proximal tubules. In particular, ninety percent of glucose reuptake in the kidney occurs in the epithelial cells of the early S1 segment of the renal cortical proximal tubule. Sodium-dependent glucose transporter 2 (SGLT2) appears to be the major transporter responsible for the reuptake of glucose at this site (Kanai et al., (1994) J Clin Investig 93: 397-404).
SGLT2 is a 672 amino acid protein containing 14 membrane-spanning segments that is predominantly expressed in the early S1 segment of the renal proximal tubules. The substrate specificity, sodium-dependence, and localization of SGLT2 are consistent with the properties of the high capacity, low affinity, sodium-dependent glucose transporter previously characterized in human cortical kidney proximal tubules. In addition, hybrid depletion studies in rats implicated SGLT2 as the predominant Na+/glucose cotransporter in the S1 segment of the proximal tubule, since virtually all sodium-dependent glucose transport activity was inhibited by an antisense oligonucleotide specific to rat SGLT2 (You et al., (1995) J Biol Chem. 270(49):29365-71).
SGLT2 is also a candidate gene for some forms of familial glucosuria, a genetic abnormality in which renal glucose reabsorption is impaired to varying degrees (van den Heuvel et al., (2002) Hum Genet 111: 544-547; and Calado et al., (2004) Hum Genet 114: 314-316). The familial glycosuria syndromes are conditions in which intestinal glucose transport is normal and renal transport of other ions and amino acids is also normal. Familial glycosuria patients appear to develop normally, have normal plasma glucose levels, and appear to suffer no major health deficits as a consequence of their disorder, despite sometimes quite high (110-114 g/daily) levels of glucose excreted. The major symptoms evident in these patients include polyphagia, polyuria, and polydipsia. The kidneys appear to be normal in structure and function. Thus, from the evidence available so far, defects in renal reuptake of glucose appear to have minimal long term negative consequences in otherwise normal individuals. Studies of highly homologous rodent SGLTs also strongly implicate SGLT2 as the major renal sodium-dependent transporter of glucose and suggest that SGLT2 activity plays some role in glucosuria.
SGLT1, another sodium-dependent glucose cotransporter that is 60% identical to SGLT2 at the amino acid level, is expressed in the small intestine and in the more distal S3 segment of the renal proximal tubule (Pajor and Wright (1992) J Biol Chem 267: 3557-3560; and Wright (2001) Am J Physiol 280: F10-F18). Despite their sequence similarities, human SGLT1 and SGLT2 are biochemically distinguishable. For SGLT1, the molar ratio of Na+ to glucose transported is 2:1. For SGLT2, the ratio is 1:1. The Km for Na+ is 32 mM for SGLT1 and 250-300 mM for SGLT2. SGLT1 and SGLT2 also vary in their substrate specificities for some sugars, but the Km values for the uptake of glucose and the nonmetabolizable glucose analog, alpha-methylglucoside (AMG), are similar. For glucose, the Km values are 0.8 mM and 1.6 mM for SGLT1 and SGLT2, respectively. For AMG, Km values are 0.4 mM and 1.6 mM for SGLT1 and SGLT2, respectively (Kanai et al., (1994) J Clin Investig 93: 397-404; and U.S. Patent Application No. 2008/0234367).
Administration of phlorizin, a nonspecific SGLT1/SGLT2 inhibitor, provided in vivo proof of concept data for use of SGLT inhibitors to treat disorders associated with hyperglycemia (e.g., NIDDM and Syndrome X). Administration of phlorizin promoted glucose excretion, lowered fasting and fed plasma glucose levels, and promoted glucose utilization without hypoglycemic side effects in rodent models of diabetes and in one canine diabetes model (Ehrenkranz et al., (2005) Diabetes Metab Res Rev 21: 31-38). No adverse effects on plasma ion balance, renal function or renal morphology were observed as a consequence of phlorizin treatment for as long as two weeks. In addition, no hypoglycemic or other adverse effects were observed when phlorizin was administered to normal animals, despite the presence of glycosuria. Furthermore, long-term treatment with synthetic agents derived from phlorizin were reported to improve fasting and fed plasma glucose, improve insulin secretion and utilization in obese type II diabetes (NIDDM) rat models, and offset the development of nephropathy in the absence of hypoglycemic or renal side effects (Ueta et al., (2005) Life Sci. 76(23):2655-68).
Phlorizin itself, however, is unattractive as an oral drug because it is a nonspecific SGLT1/SGLT2 inhibitor and because it is hydrolyzed in the gut to the aglycone, phloretin. The hydrolyzed product is a potent inhibitor of facilitated glucose transporters (GLUTs) and concurrent inhibition of GLUTs is undesirable (Katsuno et al., (2007) J Pharmacol Exp Ther. 320(1):323-30). Such inhibitors would be predicted to exacerbate peripheral insulin resistance as well as promote hypoglycemia in the CNS. Inhibition of SGLT1 could also have serious adverse consequences as is illustrated by the hereditary syndrome glucose/galactose malabsorption (GGM), in which mutations in the SGLT1 cotransporter result in impaired glucose uptake in the intestine and life-threatening diarrhea and dehydration (Turk et al., (1991) Nature 350: 354-356; and Martin et al., (1996) Nat Genet 12: 216-220).
Taken as a whole, these data suggest that specific inhibition of SGLT2 in diabetic patients may safely normalize plasma glucose by enhancing the excretion of glucose in the urine, thereby improving insulin sensitivity and delaying the development of diabetic complications. Fortunately, the biochemical differences between SGLT2 and SGLT1, as well as the degree of sequence divergence between them, allow for identification of selective SGLT2 inhibitors. What are still needed, however, to further enable the discovery and optimization of such inhibitors, are assays to test the inhibitors in vivo and evaluate the effects of inhibitors with varying potency and selectivity for SGLT2 and SGLT1.
Glucose analogs have long been used for the study of glucose transport and for the characterization of glucose transporters (for review, see Gatley (2003) J Nucl Med. 44(7):1082-6). Alpha-methylglucoside (AMG) is often the analog of choice for cell-based assays designed to study the activity of SGLT1 and/or SGLT2.
